Astrocyte- and microglia-targeting peptides were identified and isolated using phage display technology. A series of procedures, including three cycles of both in vivo and in vitro biopanning, was performed separately in astrocytes and in M1 or M2 microglia, yielding 50–58 phage plaques in each cell type. Analyses of the sequences of this collection identified one candidate homing peptide targeting astrocytes (AS1[C-LNSSQPS-C]) and two candidate homing peptides targeting microglia (MG1[C-HHSSSAR-C] and MG2[C-NTGSPYE-C]). To determine peptide specificity for the target cell in vitro, each peptide was synthesized and introduced into the primary cultures of astrocytes or microglia. Those peptides could bind to the target cells and be selectively taken up by the corresponding cell, namely, astrocytes, M1 microglia, or M2 microglia. To confirm cell-specific gene delivery to M1 microglia, the complexes between peptide MG1 and siRNA-interferon regulatory factor 5 were prepared and intrathecally injected into a mouse model of neuropathic pain. The complexes successfully suppressed hyperalgesia with high efficiency in this neuropathic pain model. Here, we describe a novel gene therapy for the treatment neuropathic pain, which has a high potential to be of clinical relevance. This strategy will ensure the targeted delivery of therapeutic genes while minimizing side effects to non-target tissues or cells.

使用噬菌体展示技术鉴定并分离了靶向星形胶质细胞和小胶质细胞的肽。在星形胶质细胞和M1或M2小胶质细胞中分别进行了一系列程序，包括体内和体外生物淘选的三个周期，在每种细胞类型中产生50-58个噬菌斑。该集合的序列分析确定了一种靶向星形胶质细胞的候选归巢肽（AS1 [C-LNSSQPS-C]）和两种靶向小胶质细胞的候选归巢肽（MG1 [C-HHSSSAR-C]和MG2 [C-NTGSPYE-C]） 。确定体外对靶细胞的肽特异性合成每种肽并将其引入星形胶质细胞或小胶质细胞的原代培养物中。这些肽可以与靶细胞结合，并被相应的细胞，即星形胶质细胞，M1小胶质细胞或M2小胶质细胞选择性摄取。为了确认将细胞特异性基因递送至M1小胶质细胞，制备了肽MG1和siRNA-干扰素调节因子5之间的复合物，并将其鞘内注射到神经性疼痛的小鼠模型中。在该神经性疼痛模型中，复合物成功地高效抑制了痛觉过敏。在这里，我们描述了一种新的基因疗法，用于治疗神经性疼痛，具有很高的临床意义。该策略将确保治疗性基因的靶向递送，同时最大程度地减少对非靶向组织或细胞的副作用。